Journal: BioResearch Open Access

Article Title: Constitutive Expression of Human Telomerase Enhances the Proliferation Potential of Human Mesenchymal Stem Cells

doi: 10.1089/biores.2012.0252

Figure Lengend Snippet: Phenotype and growth characteristics of human mesenchymal stem cell (hMSCs) and hMSC telomerase reverse transcriptase (TERT) cells. (A) Late-passage [P15(19)] hMSC TERT cells still exhibit the characteristic fibroblastic phenotype of early-passage (P3) hMSCs. (B) Mock-transduced hMSCs and hMSC TERT cells were serial passaged in cell culture, and the population-doubling level (PDL) was calculated at each passage number. At early passages [P2(6)–P7(11)], the PDLs are similar for the two cell lines, but after P8(12), hMSCs proliferate at a slower rate and ceased growth at P16(20) with a maximum PDL of 14. hMSC TERT cells continually proliferate even after PDL 70 [P40(44)].

Article Snippet: hMSCs, hMSC complete growth medium (HMSCGM), and growth supplements were obtained from Lonza, Inc. (Allendale, NJ). hMSCs used in this study were from a 32-year-old Caucasian man (Lonza Lot # 4F0591).

Techniques: Reverse Transcription, Cell Culture

Journal: BioResearch Open Access

Article Title: Constitutive Expression of Human Telomerase Enhances the Proliferation Potential of Human Mesenchymal Stem Cells

doi: 10.1089/biores.2012.0252

Figure Lengend Snippet: TERT expression in hMSCs and hMSC TERT cells. (A) Relative changes in TERT mRNA expression were determined by real-time reverse transcription (RT)–polymerase chain reaction (PCR) and expressed in relation to 18S levels, and the reference point [mock-transduced hMSC P4(8) levels] using the 2 −ΔΔC(T) method. hMSCs have a low, but detectable, level of TERT mRNA expression at P4(8), but with continued passage TERT expression quickly decreases to nearly undetectable levels even by early-passage P6(10). hMSC TERT cells have significantly high levels of TERT mRNA at all passages, even late passage P16(20). * p <0.05 compared to hMSC levels at each passage. (B) TERT enzymatic activity was also measure at both early [P6(10)] and late [P16(20) and P26(30)] passages. There are significantly higher levels of TERT activity in hMSC TERT cell extracts than in hMSC extracts throughout the time course of the study. * p <0.05 compared to hMSC levels at P6(10).

Article Snippet: hMSCs, hMSC complete growth medium (HMSCGM), and growth supplements were obtained from Lonza, Inc. (Allendale, NJ). hMSCs used in this study were from a 32-year-old Caucasian man (Lonza Lot # 4F0591).

Techniques: Expressing, Reverse Transcription, Polymerase Chain Reaction, Activity Assay

Journal: BioResearch Open Access

Article Title: Constitutive Expression of Human Telomerase Enhances the Proliferation Potential of Human Mesenchymal Stem Cells

doi: 10.1089/biores.2012.0252

Figure Lengend Snippet: Expression of p16 INK4a cell cycle regulator in hMSCs and hMSC TERT cells. mRNA levels were quantitated by real-time RT-PCR and normalized to expression levels at P6(10) for each cell line. p16 INK4a levels are significantly increased in hMSCs at later passages [P13(17)], indicating aging of the culture. Levels in hMSC TERT cell lines are unchanged. * p <0.05 compared to P6(10) levels for each cell line.

Article Snippet: hMSCs, hMSC complete growth medium (HMSCGM), and growth supplements were obtained from Lonza, Inc. (Allendale, NJ). hMSCs used in this study were from a 32-year-old Caucasian man (Lonza Lot # 4F0591).

Techniques: Expressing, Quantitative RT-PCR

Journal: BioResearch Open Access

Article Title: Constitutive Expression of Human Telomerase Enhances the Proliferation Potential of Human Mesenchymal Stem Cells

doi: 10.1089/biores.2012.0252

Figure Lengend Snippet: Enhanced growth characteristics of hMSC TERT cells due to increased cell proliferation and decreased cellular senescence. (A) Cellular proliferation was assayed by measurement of BrdU incorporation during DNA replication, indicating that at both early [P8(12)] and late [P16(20)] passage, the proliferative rate of hMSC TERT cells is significantly higher than that of hMSCs (mock-transduced). * p <0.05 compared to hMSC levels at each passage. The proliferative rate of hMSCs decreases significantly from P8(12) to P16(20), whereas the rate of proliferation of hMSC TERT cells decreases slightly but insignificantly over the same passages. § p <0.05 compared to hMSC levels at P8(12). (B) Cellular senescence was measured by the histochemical stain for β-galactosidase activity. There is a significant increase in β-galactosidase-positive cells in hMSCs after passage P12, whereas the percentage of positive hMSC TERT cells did not increase even after P40(44). * p <0.05 compared to hMSC levels at passage P10. (C) Apoptosis potential was determined by colorimetric determination of DNA fragmentation in hMSCs or hMSC TERT cells. There was no significant difference in the level of apoptosis, although the trend was for lower apoptosis in hMSC TERT cells in comparison to mock-transduced hMSCs.

Article Snippet: hMSCs, hMSC complete growth medium (HMSCGM), and growth supplements were obtained from Lonza, Inc. (Allendale, NJ). hMSCs used in this study were from a 32-year-old Caucasian man (Lonza Lot # 4F0591).

Techniques: BrdU Incorporation Assay, Staining, Activity Assay, Comparison

Journal: BioResearch Open Access

Article Title: Constitutive Expression of Human Telomerase Enhances the Proliferation Potential of Human Mesenchymal Stem Cells

doi: 10.1089/biores.2012.0252

Figure Lengend Snippet: Chemotaxis toward a medium containing 1% fetal bovine serum (FBS). Data are expressed as a chemotactic index, defined as the number of cells migrating in response to the medium (1% FBS) divided by the number of cells migrating in the negative control (−). hMSC TERT cells [P12(16)] exhibit significantly higher levels of chemotaxis than hMSCs at earlier PDL (P8). * p <0.05 compared to negative control.

Article Snippet: hMSCs, hMSC complete growth medium (HMSCGM), and growth supplements were obtained from Lonza, Inc. (Allendale, NJ). hMSCs used in this study were from a 32-year-old Caucasian man (Lonza Lot # 4F0591).

Techniques: Chemotaxis Assay, Negative Control